Brain catalase in the streptozotocin-rat model of sporadic Alzheimer's disease treated with the iron chelator-monoamine oxidase inhibitor, M30.

Low intracerebroventricular (icv) doses of streptozotocin (STZ) produce regionally specific brain neurochemical changes in rats that are similar to those found in the brain of patients with sporadic Alzheimer's disease (sAD). Since oxidative stress is thought to be one of the major pathologic processes in sAD, catalase (CAT) activity was estimated in the regional brain tissue of animals treated intracerebroventricularly with STZ and the multitarget iron chelator, antioxidant and MAO-inhibitor M30 [5-(N-methyl-N-propargylaminomethyl)-8-hydroxyquinoline]. Five-day oral pre-treatment of adult male Wistar rats with 10 mg/kg/day M30 dose was followed by a single injection of STZ (1 mg/kg, icv). CAT activity was measured colorimetrically in the hippocampus (HPC), brain stem (BS) and cerebellum (CB) of the control, STZ-, M30- and STZ + M30-treated rats, respectively, 4 weeks after the STZ treatment. STZ-treated rats demonstrated significantly lower CAT activity in all three brain regions in comparison to the controls (p < 0.05 for BS and CB, p < 0.01 for HPC). M30 pre-treatment of the control rats did not influence the CAT activity in HPC and CB, but significantly increased it in BS (p < 0.05). M30 pre-treatment of STZ-treated rats significantly increased CAT activity in the HPC in comparison to the STZ treatment alone (p < 0.05) and normalized to the control values. These findings are in line with the assumption that reactive oxygen species contribute to the pathogenesis of STZ in a rat model of sAD and indicate that multifunctional iron chelators such as M30 might also have beneficial effects in this non-transgenic sAD model.

Genomic aspects of sporadic Parkinson's disease.

Parkinson's disease (PD) is thought to be associated with oxidative stress mechanisms, as well as with glutamate receptor abnormalities, ubiquitin-proteasome dysfunction, inflammatory and cytokine activation, dysfunction in neurotrophic factors, damage to mitochondria, cytoskeletal abnormalities, synaptic dysfunction and activation of apoptotic pathways. To investigate these hypotheses, many researchers have applied molecular biology techniques to the study of neuronal cell death in these conditions. In this article, we discuss recent findings of gene expression in PD that may elucidate the usage of specific new biomarkers for sporadic PD and point to novel drug developments.

Induction of meiosis in Saccharomyces cerevisiae depends on conversion of the transcriptional represssor Ume6 to a positive regulator by its regulated association with the transcriptional activator Ime1.

The transcription of meiosis-specific genes, as well as the initiation of meiosis, in the budding yeast Saccharomyces cerevisiae depends on IME1. IME1 encodes a transcriptional activator which lacks known DNA binding motifs. In this study we have determined the mode by which Ime1 specifically activates the transcription of meiotic genes. We demonstrate that Ime1 is recruited to the promoters of meiotic genes by interacting with a DNA-binding protein, Ume6. This association between Ime1 and Ume6 depends on both starvation and the activity of a protein kinase, encoded by RIM11 In the absence of Ime1, Ume6 represses the transcription of meiotic genes. However, in the presence of Ime1, or when Ume6 is fused in frame to the Gal4 activation domain, Ume6 is converted from a repressor to an activator, resulting in the transcription of meiosis-specific genes and the formation of asci.

Green tea catechins as brain-permeable, non toxic iron chelators to "iron out iron" from the brain.

Evidence to link abnormal metal (iron, copper and zinc) metabolism and handling with Parkinson's and Alzheimer's diseases pathology has frequently been reported. The capacity of free iron to enhance and promote the generation of toxic reactive oxygen radicals has been discussed numerous times. Metal chelation has the potential to prevent iron-induced oxidative stress and aggregation of alpha-synuclein and beta-amyloid peptides. The efficacy of iron chelators depends on their ability to penetrate the subcellular compartments and cellular membranes where iron dependent free radicals are generated. Thus, natural, non-toxic, brain permeable neuroprotective drugs, are preferentially advocated for "ironing out iron" from those brain areas where it preferentially accumulates in neurodegenerative diseases. This review will discuss the most recent findings from in vivo and in vitro studies concerning the transitional metal (iron and copper) chelating property of green tea, and its major polyphenol, (-)-epigallocatechin-3-gallate with respect to their potential for the treatment of neurodegenerative diseases.

Involvement of multiple survival signal transduction pathways in the neuroprotective, neurorescue and APP processing activity of rasagiline and its propargyl moiety.

Our recent studies aimed to elucidate the molecular and biochemical mechanism of actions of the novel anti-Parkinson's drug, rasagiline, an irreversible and selective monoamine oxidase (MAO)-B inhibitor and its propargyl moiety, propargylamine. In cell death models induced by serum withdrawal in rat PC12 cells and human SH-SY5Y neuroblastoma cells, both rasagiline and propargylamine exerted neuroprotective and neurorescue activities via multiple survival pathways, including: stimulation of protein kinase C (PKC) phosphorylation; up-regulation of protein and gene levels of PKCalpha, PKCepsilon and the anti-apoptotic Bcl-2, Bcl-xL, and Bcl-w; and up-regulation of the neurotrophic factors, BDNF and GDNF mRNAs. Rasagiline and propargylamine inhibited the cleavage and subsequent activation of pro-caspase-3 and poly ADP-ribose polymerase. Additionally, these compounds significantly down-regulated PKCgamma mRNA and decreased the level of the pro-apoptotic proteins, Bax, Bad, Bim and H2A.X. Rasagiline and propargylamine both regulated amyloid precursor protein (APP) processing towards the non-amyloidogenic pathway. These structure-activity studies have provided evidence that propargylamine promoted neuronal survival via neuroprotective/neurorescue pathways similar to that of rasagiline. In addition, recent study demonstrated that chronic low doses of rasagiline administered to mice subsequently to 1 methyl-4 phenyl 1,2,3,6 tetrahydropyridine (MPTP), rescued dopaminergic neurons in the substantia nigra pars compacta via activation of the Ras-PI3K-Akt survival pathway, suggesting that rasagiline may possess a disease modifying activity.

The triangle inequality in balanced genetical polymorphisms.

A simplified derivation of the "triangle inequality," first obtained by Lewontin, Ginzburg and Tuljapurkar (1978), is presented.

Recurrent rhabdomyolysis as a manifestation of alcoholic myopathy. Report of a case.

A 38-year-old man experienced six severe episodes of rhabdomyolysis and two episodes of pharyngeal muscle weakness superimposed on chronic alcoholic myopathy and complicated by cardiomyopathy. A muscle biopsy specimen demonstrated sharply reduced levels of electrolytes despite normal serum values; presumably, these deficiencies were related to the pathogenesis of the recurrent rhabdomyolysis.

Thyroiditis after pregnancy loss.

We present a case series evaluating the development and characteristics of thyroiditis following pregnancy loss. Five women were followed prospectively with measurement of thyroid function and antithyroid antibodies after pregnancy loss. Serum TSH concentrations were measured by immunoradiometric assay and antithyroid antibodies by RIA and hemagglutination techniques. All women had normal serum TSH concentrations before conception or at the time of pregnancy loss, and all but one had positive antithyroid antibodies. Pregnancy loss occurred between 5-20 weeks gestation because of ectopic pregnancy or either spontaneous or elective abortion. Two women had subclinical hypothyroidism with peak serum TSH values of 8.7 mU/L and 5.4 mU/L at 2 and 7 months after pregnancy loss, respectively. Three women had clinical hyperthyroidism with serum TSH values < or = 0.2 mU/L diagnosed between 3-11 months after pregnancy loss followed subsequently by a hypothyroid phase. Painless thyroiditis within 1 yr of pregnancy loss in these women suggests that the immunological changes of a short-term gestation may be sufficient to lead to thyroiditis.

Activating mutation in the stimulatory guanine nucleotide-binding protein in an infant with Cushing's syndrome and nodular adrenal hyperplasia.

Cushing's syndrome in infancy is uncommon. In this report, we describe an infant with ACTH-independent Cushing's syndrome in which an activating mutation in the stimulatory G-protein (Gs alpha) was detected. The patient presented at 3 months of age with Cushingoid features, poor linear growth, and elevated liver enzymes. Plasma ACTH and dexamethasone suppression test results were consistent with ACTH-independent Cushing's syndrome, and a subsequent adrenalectomy revealed bilateral adrenocorticonodular hyperplasia. Asymptomatic lesions consistent with fibrous dysplasia were later detected on bone scan. Genomic DNA was extracted from adrenal, liver, and blood and amplified by polymerase chain reaction with Gs alpha exon 8 primers. Using allele-specific oligonucleotide hybridization, the DNA was probed for known Gs alpha-activating mutations. A point mutation coding for an arginine to cysteine substitution at codon 201 of exon 8 was detected in genomic DNA from this infant's adrenal, liver, and leukocytes. The mutation was detected in nodular adrenal tissue, but was essentially absent in normal adrenal tissue. Activating mutations in the Gs alpha gene have previously been described in GH-secreting tumors, thyroid adenomas, and the McCune-Albright syndrome and are probably involved in the pathogenesis of adrenocorticonodular hyperplasia in this infant with Cushing's syndrome.

Changes in insulin-like growth factor-I (IGF-I), IGF-binding protein-3, growth hormone (GH)-binding protein, erythrocyte IGF-I receptors, and growth rate during GH treatment.

To assess the relative determinants of growth rate, we measured serum levels of insulin-like growth factor-I (IGF-I), IGF-binding protein-3 (IGFBP-3), and GH-binding protein (GHBP) as well as IGF-I erythrocyte receptor specific binding (SB) in 14 prepubertal GH-deficient children before and during the first year of treatment with 0.043 mg/kg.day GH. Serum IGF-I and IGFBP-3 levels, measured by RIA, were significantly increased by 2 weeks and showed progressive increases throughout the year of GH therapy. Growth rate (height velocity SD score adjusted for bone age) correlated best with the 12 month changes in IGFBP-3 (r = 0.81; P < 0.001) and IGF-I (r = 0.72; P = 0.005), and to a lesser extent with the 12 month absolute IGFBP-3 (r = 0.58; P = 0.04) and the 6 month change in IGFBP-3 (r = 0.55; P = 0.05). The baseline IGF-I correlated inversely with the growth rate during GH therapy (r = -0.55; P = 0.05) and was the best pretreatment predictor of growth response. GHBP, as measured by ligand-mediated immunofunctional assay, showed no significant change during GH therapy and did not correlate with growth response. The baseline GHBP, however, did correlate with both the 12 month IGFBP-3 (r = 0.72; P = 0.006) as well as the 2 week change in IGFBP-3 (r = 0.63; P = 0.05). Erythrocyte IGF-I SB showed a significant decrease by 6 months secondary to a decrease in IGF-I receptor number, with no change in affinity. The 6 month IGF-I receptor binding correlated inversely with the increase in IGF-I (r = -0.88; P < 0.001). Erythrocyte IGF-I SB at baseline did not correlate with the growth response, although there was an inverse trend between the 6 month IGF-I receptor level and the growth rate. IGF-I and IGFBP-3 show progressive increases, whereas the erythrocyte IGF-I receptor-binding capacity decreases by 6 months, and GHBP shows little change during the first year of GH treatment. Data from this study suggest that changes in IGFBP-3 and, to a lesser extent, IGF-I are the major correlates of growth rate, and that down-regulation of the IGF-I receptor may have relatively little influence on growth rate compared with changes in IGFBP-3 and IGF-I.

Interrelationships between the renin-angiotensin-aldosterone and calcium homeostatic systems.

Blood pressure is affected by both sodium and calcium intake. To determine if there is an interaction between the regulatory mechanisms for these two cations, eight normal male volunteers received the following 1-h infusions on three different days: 1) angiotensin II (AII), 2) the synthetic 1-34 amino terminal fragment of human PTH [hPTH(1-34)], and 3) AII and hPTH(1-34) together. Blood samples were obtained at t = 0 and every 20 min during each infusion and urine was collected for 3 h both before and after the start of each infusion. Infusion of AII produced an increase in intact PTH from 18 +/- 2 to 31 +/- 4 ng/L (P < 0.05), most likely in response to a small decrease in serum ionized calcium (1.25 +/- 0.01 to 1.23 +/- 0.01 mmol/L, P < 0.05). Urinary excretion of calcium was unchanged. Infusion of hPTH(1-34) at 200 U/h increased N-terminal PTH levels (18 +/- 3 to 268 +/- 42 ng/L, P < 0.05), decreased tubular reabsorption of phosphate (0.92 +/- 0.03 to 0.82 +/- 0.11, P < 0.05), and increased urinary cAMP (0.18 +/- 0.02 to 0.53 +/- 0.05 nmol/L of glomerular filtrate, P = 0.0001). hPTH(1-34) infusion suppressed endogenous intact PTH (18 +/- 3 to 14 +/- 2 ng/L, P < 0.005) and increased PRA from 0.14 +/- 0.02 to 0.32 +/- 0.05 ng/(L.s) (P < 0.05) without a change in serum ionized calcium which suggests direct effects of hPTH(1-34) on the parathyroid glands and the juxtaglomerular apparatus. The effects of AII and hPTH(1-34) were antagonistic with little change in serum ionized calcium, intact PTH, or PRA when both were infused together. These interrelationships between the major hormonal systems controlling sodium and calcium homeostasis suggest a mechanism underlying the close association of calcium and sodium in the regulation of blood pressure.

The hypothalamic-pituitary-thyroid negative feedback control axis in children with treated congenital hypothyroidism.

Measurements of serum concentrations of free T4, T3, TSH, and thyroglobulin (Tg) were conducted in 42 infants (2-9 months of age) detected and treated through the Northwest Newborn Regional Screening Program and 63 children and adolescents (1-18 yr of age) with congenital hypothyroidism (CH) detected and managed in the Northern California Kaiser Permanente Medical Care Program. Normal feedback control axis data were developed by Quest Diagnostics, Inc. - Nichols Institute Diagnostics and Loma Linda University, from free T4 and TSH measurements in 589 healthy subjects, 2 months to 54 yr of age; 83 untreated hypothyroid patients; and 116 untreated hyperthyroid patients. Twenty-four of the 42 CH infants and 57 of the 63 CH children manifested serum TSH concentrations appropriate for the measured free T4 level. In the remaining 18 infants and 6 children, serum free T4 values were increased 0.2-1.4 ng/dL (2.6-18.0 pmol/L) for the prevailing TSH level, suggesting a state of mild to moderate pituitary-thyroid hormone resistance. In the treated children, the mean T3 concentration was lower (by 32%, 102 vs. 150 ng/dL; 1.57 vs. 2.31 nmol/L) than in normal children, in agreement with earlier data in hypothyroid adults treated with exogenous T4. Serum Tg concentrations were normal or elevated in 90% of the 19 children with ectopic glands and 93% of 27 children with eutopic glands in whom measurements were available. There was a positive correlation between serum TSH and Tg concentrations (P < 0.001), suggesting significant endogenous thyroid hormone production in these children. Our results suggest that the majority of infants and children with CH have a normal hypothalamic-pituitary-thyroid negative feedback control axis during treatment and that the measurement of serum TSH is a useful marker complementing the free T4 measurement in the management of children with CH. A minority have variable pituitary-thyroid hormone resistance, with relatively elevated serum TSH levels for their prevailing serum free T4 concentration. The prevalence of resistance is greater (43%) in young infants (< 1 yr of age) than in older children (10%), indicating that, in most children, the resistance improves with age.

Cloning and in vitro expression of the human selenoprotein, type I iodothyronine deiodinase.

The type I 5' iodothyronine deiodinase (5' DI) catalyzes the deiodination of T4 to the biologically active hormone T3 and accounts for a significant fraction of its production. We have recently cloned the complementary DNA (cDNA) for the rat 5' DI, which contains the rare amino acid selenocysteine, and used this to screen human liver and kidney cDNA libraries to identify a human 5' DI cDNA clone. From these, we constructed a cDNA encoding a functional 5' DI. The 2222 base pair human 5' DI cDNA is approximately 200 nucleotides shorter than the 2.4-kilobase hybridizing band in Northern blots of human liver, kidney, and thyroid, because of missing 5' untranslated sequence and the poly A tail. The deduced amino acid sequence codes for a protein of 28.7 kilodaltons assuming the UGA codon at position 382 encodes selenocysteine, and is highly homologous (88% similarity) to the rat. We transiently expressed the 5' DI in COS-7 cells to establish that it encodes a functional enzyme and to study its kinetics. These show saturable deiodination of rT3 (Ka 0.52 +/- 0.04 mumol/L and Vmax 63.2 +/- 16.4 pmol min-1 mg-1). T4 and gold thioglucose are competitive inhibitors of rT3 deiodination. 6-n-Propylthiouracil (PTU) is an uncompetitive inhibitor (with rT3) and competitive inhibitor (with dithiothreitol) of rT3 deiodination. 6-n-Propylthiouracil inhibits T4 to T3 conversion. Labeling of COS-7 cells transiently transfected with the human 5' DI cDNA with bromoacetyl-125I-T3 demonstrates a 28-kilodalton protein. This indicates that in the human, as well as in the rat messenger RNA, the UGA encodes selenocysteine and translation terminates at the UAA codon at nucleotides 754 to 756. Reverse T3 and gold thioglucose (100 nmol/L) block bromoacetyl-125I-T3 labeling of the transiently expressed human and rat 5' DI proteins. These results demonstrate that the human 5' DI is a selenoprotein, analogous to the rat enzyme. Given the previously demonstrated critical role of the selenium atom in catalyzing deiodination by this protein, we conclude that this trace element is essential for normal thyroid hormone action in man.

Hypothalamic pituitary adrenal function in the extremely low birth weight infant.

Extremely premature infants manifest clinical features suggestive of adrenal insufficiency. Yet, serum cortisol levels are similar in ill and well preterm infants in a setting where one would expect high stress levels in the ill infants. We investigated the hypothalamic-pituitary-adrenal axis in 17 extremely low birth weight stressed premature infants, mean birth weight 739 g, gestational age, 26.1 weeks, using ovine CRH (oCRH) and ACTH stimulation. oCRH (1 microgram/kg) was administered at 2-7 days of life (mean = 4.1). ACTH rose from a basal value 6.0 +/- 0.8 pmol/L (mean +/- SEM) to 9.6 +/- 1.8 pmol/L (P < 0.01) at 15 min and 9.5 +/- 1.7 pmol/L (P < 0.01) at 60 min. Basal cortisol rose from 349.3 +/- 58.1 nmol/L to 422.3 +/- 57.9 nmol/L (P < 0.01) at 15 min and 568.7 +/- 60.2 nmol/L (P < 0.01) at 60 min. Cortisol values remained significantly (P < 0.05) elevated 24 h after oCRH. An ACTH stimulation test performed 24 h after the oCRH test demonstrated a significant cortisol rise from 603.5 +/- 130.5 nmol/L to 882.7 +/- 136.6 nmol/L (P < 0.05) at 60 min. Plasma CRH immunoactivity was also measured before oCRH testing and was detectable in 10 of 15 infants. The mean CRH immunoactivity was 21.8 +/- 4.4 pmol/L in the infants, significantly higher than 8 adult male controls (P < 0.04). Our results show a normal pituitary response to ovine CRH and a normal adrenal response to ACTH. We hypothesize that cortisol levels are inappropriately low in some ill preterm infants because of the inability of the extremely premature brain to recognize the stress of the illness or because of inadequate hypothalamic secretion of CRH. The significance of the measurable plasma CRH in the first week of life is unknown.

Alterations in body weight and composition consequent to 20 wk of endurance training: the HERITAGE Family Study.

BACKGROUND: Obesity is a major public health problem in the United States. The role of physical activity and formal exercise in controlling body weight has not been clearly determined. OBJECTIVE: This study determined the magnitude of change in body weight and composition across sex, race, and age in response to 20 wk of endurance training. DESIGN: Men and women (n = 557) of various ages (16-65 y) and 2 races (black and white) exercised on cycle ergometers 3 d/wk for a total of 60 exercise sessions starting at 55% of maximal oxygen consumption (VO(2)max) for 30 min/session and building to 75% of VO(2)max for 50 min/session, where it was maintained during the last 6 wk. Skinfold-thickness measurements, circumferences, body composition (by hydrostatic weighing), and body fat distribution (by computed tomography scan at L4-L5 and the waist-hip ratio) were determined before and after training. RESULTS: All skinfold-thickness and circumference measures, waist-hip ratio, body mass index, total body mass, fat mass, percentage body fat, and computed tomography scan measures of total, subcutaneous, and visceral abdominal fat decreased with training, whereas total body density and fat-free mass increased. These changes were significant, but small. There were several differences in training response by sex and race, but not by age. CONCLUSIONS: A short-term exercise intervention can induce favorable changes in body composition, but the magnitude of these changes is of limited biological significance. Increasing physical activity likely has a major effect on body-composition and fat distribution characteristics only when it is of a greater magnitude and sustained for much longer periods

Wound zygomycosis (mucormycosis) in otherwise healthy adults.

Two previously healthy men sustained trauma that caused extensive soft tissue damage together with soil contamination. Within three days, rapidly advancing necrosis was observed at the wound margins. Histologic examination revealed the presence of non-septate branching hyphae characteristic of Mucorales within tissues and in the lumen of blood vessels. In one case, the disease was unrecognized until widespread dissemination had taken place, and the patient died; in the other, a cure resulted from aggressive medical and surgical management. Infections due to Mucorales generally occur in immune-compromised hosts. In cases of extensive trauma, inoculation of devitalized tissues with soil may initiate infection by zygomycetes, even in persons whose immunologic status appears to be normal.

Constitutional delay of growth: expected versus final adult height.

Constitutional delay of growth and puberty is believed to represent a variation of normal growth, and it is expected that children with this condition will grow for a longer duration than average and reach a height that is normal for their genetic potential. The records of children with constitutional delay of growth and puberty who were initially seen in the Pediatric Endocrine Clinic at the Oregon Health Sciences University between 1975 and 1983 were retrospectively reviewed. Criteria for study included a height more than 2 SD below the mean, a significantly delayed bone age, and a normal growth velocity on follow-up. Forty-two subjects were located and final adult height measurements were obtained. AT contact, the 29 male subjects (mean age = 23.9 years) were 169.5 +/- 4.5 cm tall (mean +/- SD), and the 13 female subjects (mean age = 20.5 years) were 156 +/- 3.8 cm tall. Adult height predictions during follow-up, using either the Bayley-Pinneau or Roche-Wainer-Thissen method, were close to final adult heights. The males were 1.2 SD and the females 1.3 SD below the 50th percentile as adults. This finding was not fully explained by genetic short stature; the males fell 5.1 cm and the females 5.3 cm below target heights based on midparental heights. It is concluded that this discrepancy is most likely explained by a selection bias of the shortest children referred to and observed in a subspecialty clinic, although a defect in human growth hormone secretion or function in children at the far end of the spectrum of constitutional delay of growth and puberty cannot be excluded.

Thrombosis: a study of coagulation parameters and mechanisms during allograft rejection.

Thrombosis of the microvasculature has been recognized at the end product of organ rejection, but the exact biological pathway through which this occurs has not been clarified. Normal, factor VII deficient and heterozygous hemophilic (factor VIII) dogs were grouped to study the intrinsic and extrinsic clotting and platelet mechanisms during unmodified renal allograft rejection. The observed alterations of the hemostatic mechanisms are related to the changes observed in the microvasculature. Six groups of donor-recipient animals were studied: Group I -autografts (control); Group II - normal to normal allografts with bilateral nephrectomy; Group III - heterozygotes for factor VIII deficiency; Group IV - normal to normal allografts with unilateral nephrectomy; Group V - normal to factor VII deficiency without nephrectomy; and Group VI - normal to normal allografts with unilateral nephrectomy and dipyridamole. Each engrafted animal was followed pre- and posttransplantation for change in the blood clotting factors, fibrin split products, platelets, white blood cells, renal function and microvasculature. The animals with factor VII deficiency rejected in a similar fashion as the control animals. The group with impaired factor VIII synthesis and platelet function had longer survival times. These data suggest that the intrinsic clotting pathway and platelets are the primary mechanism through which thrombosis occurs secondary to immune injury.

Placental protein 13 (PP-13): effects on cultured trophoblasts, and its detection in human body fluids in normal and pathological pregnancies.

Placental tissue protein 13 (PP-13), one of the 56 known placental proteins identified till today, was purified from placentas obtained from women at delivery, and used to evoke antibodies against it. The purified PP-13 was lysed to peptides, which were sequenced, leading to the full-length cDNA sequencing and its expression in Escherichia coli. Sequence analysis in databases showed homology to the galectin family. Of the various antibody preparations developed, a pair of monoclonal antibodies (MAbs) coupled to the recombinant PP-13 (PP-13-R) was used for the immunodetection of PP-13 in pregnant women's serum with the solid-phase ELISA format. With a dynamic range of 25-500 pg/mL with no background in non-pregnant women's serum and men's serum, the ELISA test was suitable for the detection of PP-13 in the 1st, 2nd, and 3rd trimesters. PP-13 levels slowly increase during pregnancy. In the 1st trimester, lower than normal PP-13 levels were found in fetal growth restriction (IUGR), preeclampsia (PE), and particularly in early PE (<34 weeks of gestation). In the 2nd and 3rd trimesters, higher than normal concentrations were found in PE, IUGR and in preterm delivery (PTD). Application of PP-13 to cultured trophoblasts elicited depolarization carried by calcium ions, followed by liberation of linoleic and arachidonic acids from the trophoblast membrane, and a subsequent elevation of prostacyclin and thromboxane. These effects were negligible when PP-13 derived from the placentas of patients with IUGR, PE or PTD was used. The results are discussed in view of the potential utilization of PP-13 for early serum screening to assess the risk to develop placental insufficiency, coupled to a differential analysis of the various pathologies by analyzing cultured trophoblasts.

Prolactin and antiprolactin receptor antibody inhibit steroidogenesis by purified rat Leydig cells in culture.

The in vitro effects of ovine PRL (oPRL) on testicular testosterone synthesis were determined using isolated, collagenase-dispersed, adult rat Leydig cells in culture. oPRL (50-1000 ng/ml) had no effect either on basal or on LH (50, 100 or 2000 pg/ml)-stimulated testosterone secretion by Leydig cells in short-term culture (4 h). 125I-oPRL binding studies revealed a single class of high affinity sites (Ka 8.7 nM) with a low capacity (Bmax 6.7 fmol/mg protein identical to approximately 980 sites/Leydig cell). Isolated Leydig cells were further purified on a continuous Percoll gradient and cultured in serum-free medium, at 34 degrees C, in 5% CO2 and 95% air. After 3 days of culture, the media were collected, the cells washed and then stimulated with hCG (3 ng/ml) for 3 h. oPRL (1-1000 ng/ml) added at plating, caused a log dose-dependent inhibition of testosterone accumulation during the 3-day culture period; the highest and most consistent inhibition (31%) was with 500 ng/ml oPRL. hCG increased the sensitivity to the inhibitory effect of PRL, 10 ng/ml oPRL causing 40% inhibition and 100 ng/ml causing a maximal inhibition of 50%. PRL in fact caused a reduction in the maximal effect (efficacy) of hCG on steroidogenesis, without significantly affecting the ED50 (sensitivity). The effects of an antiPRL receptor antibody raised by the antiidiotypic route and previously shown to bind to rat testis PRL receptors were tested. The antiPRL receptor IgG (13 micrograms/ml) mimicked the PRL inhibitory effect and acted synergistically with PRL (100 ng/ml) in inhibiting both testosterone accumulation in 3-day cultured Leydig cells and their subsequent response to hCG. In summary, a clear inhibitory effect of PRL and a synergistic effect of antiPRL receptor antibody were demonstrated on testosterone synthesis by rat Leydig cells in 3-day culture.